The discovery that Cyclosporin-A (CsA) supplemented with corticosteroids could mostly control acute rejection made the transplantation of liver, lung and hearts successful, life-saving procedures. Nonetheless CsA- based therapy cannot prevent all cases of graft rejection. Acute rejection that is resistant to CsA is usually reversible through the use of high-dose corticosteroids or anti-lymphocyte sera. However a significant proportion of grafts that escape acute rejection eventually succumb to chronic rejection that cannot be controlled by currently available immunosuppressive drugs. The inability of CsA to control chronic rejection and its considerable nephrotoxicity have provided the impetus for the development of new immunosuppressive agents. Most studies have continued to focus on the ability these agents to interfere with T cell function and the ability of these agents to block B cell function is less well characterized. Leflunomide is a novel immuno-modulatory drug that profoundly suppresses cellular and humoral immune responses in vivo. It is an isoxazol derivative that has no chemical relationship to any other immunosuppressive drug. It prevents and reverses acute rejection of skin, kidney, heart and small bowel allografts in rats, and clears mononuclear cell infiltration in rejecting allografts. Leflunomide also prevents the development of allo- and antigen-specific antibodies in vivo and shuts down on-going antibody responses in mice and rats. The ability to shut down an on-going antibody response in vivo, even in the presence of sensitizing allograft or antigen, has not been reported for any other agents, and is thus unique to leflunomide. These observations raise the possibility that leflunomide may be useful in preventing chronic rejection by interfering with the development of or suppressing on-going production of allo-specific antibodies. Studies proposed here focus on defining the mechanism of leflunomide immunosuppression. Preliminary studies indicate that leflunomide inhibits T cell proliferation by interfering with responsiveness to IL-2, and that leflunomide can inhibit tyrosine kinase activity. The first specific aim is to test the hypothesis that leflunomide prevent T cell proliferation by inhibiting tyrosine kinases involved in IL-2 signal transduction. The tyrosine kinases and their phosphorylated substrates involved in IL-2-mediated signal transduction will be characterized. The second specific aim is to define the cellular and biochemical basis of leflunomide inhibition of allo-specific antibody response. the third specific aim is to determine the cellular basis for leflunomide-mediated clearing of mononuclear cell infiltration in rejecting allografts. We will test whether leflunomide alteres expression of adhesion/homing molecules on endothelium or mononuclear cells. The last specific aim is to test the ability of leflunomide to induce long-term graft acceptance and T cell anergy in vivo. It is anticipated that these studies will lead to an improved understanding of the mechanism of immunosuppression by leflunomide and thus provide a rationale for its application in clinical transplantation.